The present proposal will continue efforts in a comprehensive drug discovery group to develop HIV-1 receptor inhibitors. Such inhibitors will block the interaction of HIV-1 gp120 envelope protein with CD4 transmembrane molecule found on human T lymphocytes and monocytes. During the last funding period, human recombinant soluble CD4 molecules were produced, crystallized and the structure of the amino terminal two domains (sCD41-183) determined to 2 angstroms surface patch on the C'C" ridge of CD4 domain 1. Three sets of investigators will now exploit their collective talents in immunology structural biology and medicinal chemistry to identify lead compounds for this purpose. Project 1 will purify existing recombinant CHO-derived gp120 as well as engineer and produce baculovirus-derived gp120 from SF2 and other HIV-1 strains. In addition, truncation/deletion/glycosylation mutants of gp120 variants will be produced. These molecules will be individually complexed to sCD41-183, further purified and the complexes resolved crystallographically by the project 5. Once the structure of a single CD4-gp120 complex is determined, gp120 derived from a different strain will be selected as an additional source for complex formation and structural analysis. Comparison of CD4 binding pockets in gp120 from different viral sources will identify the common, strain-independent features of the CD4-gp120 interaction. The coordinates of the complexes will be exploited in Project 6 to generate a list of putative CD4 or gp120 binding compounds through DOCK computational methods. These will be tested for activity in a variety of directed molecular screens and HIV-1 replication assays. In addition, chemical analogs of existing, non-immunosuppressive DOCK-based organic leads (lesser than 1000 MW) will be synthesized, which were designed to bind to CD4 and are known to inhibit the HIV-1 gp120 CD4 interaction and HIV-1 viral replication at micromolar concentrations. Efforts to co-crystallize these compounds with CD4 are presently underway in Project 5 and will greatly aid the design task. Finally, direct chemical approaches to mimic the domain 1 C'C" turn as well as the C" strand will be assessed by NMR and functional studies. The non-immunosuppressive nature of potential lead compounds will be verified by functional assays in conjunction with detailed mapping studies of the class II MHC binding site on CD4.